L-amino acids are useful in many industrial fields such as pharmaceuticals, agricultural chemicals and foods. With regard to the industrially useful L-amino acid for example, there have been known L-lysine, L-threonine, L-isoleucine and L-proline used as additives for animal feeds, ingredients for health foods, amino acid transfusion, etc.; L-arginine and L-ornithine used as ingredients for hepatic function promoters, amino acid transfusion and multi-amino acid preparations; L-histidine used as hepatic function promoters and precursor for histamine; L-phenylalanine used as a precursor for sweeteners; and L-tert-leucine, L-biphenylalanine, L-cyclohexylglycine, L-dichlorophenylalanine and L-bromophenylalanine etc. used as intermediates for various pharmaceuticals. Accordingly, there has been a demand for an efficient production of those useful L-amino acids in a state of being separated from D-amino acids.
As to a process for producing the L-amino acid, there has been a conventional method where a racemic N-acylamino acid is synthesized and then only an L-form compound in the racemic modification is hydrolyzed using an enzyme called an L-aminoacylase. In result, only L-amino acid is specifically produced. As to the L-aminoacylase used for that method, there have been known, for example, an L-aminoacylase originated from Penicillium funiculosum (Patent Document 1) and an L-aminoacylase originated from Streptomyces mobaraensis (Patent Document 2).
L-aminoacylases disclosed in Patent Documents 1 and 2 are excellent in a hydrolyzing ability. Their substrate specificity is, however, still unsatisfactory whereby they are unable to recognize, as a substrate, a sterically bulky unnatural amino acid such as N-acyl-L-tert-leucine, N-acyl-L-biphenylalanine, N-acyl-L-cyclohexylglycine, N-acyl-L-dichlorophenylalanine or N-acyl-L-bromophenylalanine etc. Therefore, according to the method using L-aminoacylase disclosed in Patent Documents 1 and 2, it has been impossible to optically resolve a sterically bulky unnatural amino acid such as N-acyl-DL-tert-leucine etc. and it has been impossible to produce a sterically bulky unnatural amino acid such as L-tert-leucine which is a useful intermediate for pharmaceuticals.
The present inventors have recently found that the L-succinylaminoacylase obtained from NCA 1503 strain of Geobacillus stearothermophilus which is a kind of thermophilic bacteria is able to recognize N-succinyl-L-tert-leucine as a substrate, determined a nucleic acid sequence of the gene coding for this L-succinylaminoacylase and filed a patent application (Patent Document 3). The L-succinylaminoacylase disclosed in Patent Document 3 is able to utilize N-succinyl-L-tert-leucine as a substrate, and thus it has the substrate specificity which has not been achieved in the conventional L-aminoacylases. However, it still has a problem in terms of its enzymatic activity. Further, with regard to the substrate specificity to sterically bulky unnatural amino acids other than N-succinyl-L-tert-leucine, it is merely confirmed in Patent Document 3 that this L-succinylaminoacylase is able to utilize N-succinyl-L-cyclohexylglycine and N-succinyl-L-4-bromophenylalanine as a substrate.